Objective: To instruct the clinical medicine in Affiliated Hospital of Dali University by analyzing the resistance of Escherichia coli and Klebsiella pneumoniae from 2011 to 2012. Methods: Isolate and culture strains by using conventional methods, and identify the bacteria with VITET-2 Compact 60 automatic microbial analyzer and test the susceptibility through K-B. Results: In total 183 specimens, 146 of them were Escherichia coli: koutan 92(63%), secretions 17(11.6%), urine 16(10.9%), pus 9(6.1% 2), other 12(8.2%); 37 of them were Klebsiella pneumoniae: koutan 27(73%), secretions five(13.5%), pus 3(8%), other susceptibility test results: the resistance rates of Cefoperazone, Amikacin, Imipenem were lower, at 10.3%, 6.2%, 2.7% and 10.8%, 13.5%, 2.7%, respectively. Conclusion: The detection rate of Escherichia coli and Klebsiella pneumoniae in sputum is rising in these two years, and the resistance rates have been exceeded more than 50% of levofloxacin, cefazolin, aztreonam, ceftazidime, especially ceftazidimewas resistance rates is significantly elevated. However, resistance rates of cefoperazone, amikacin, imipenem are lower, so doctors should choose antibiotics based on susceptibility experiments.